Huntington's Disease (HD) is a debilitating autosomal dominant neurodegenerative disease, characterized by slow-onset cognitive impairment, neuropsychiatric manifestations, and loss of motor function. There is currently no cure. It is estimated that 1 in every 10,000 people in Western countries are affected or at risk for inheriting the disease from an affected parent. The genetic origin of this disease is a CAG repeat expansion in exon 1 of the huntingtin (Htt) gene. The mechanism of neurodegeneration is not fully understood, but recent studies sug- gest that transient neuronal knockdown of Htt mRNA can reverse disease progression without compromising normal cellular function in vivo. RNA interference via siRNA is the most promising therapeutic approach for transient Htt mRNA silencing. Treatment of HD and other neurodegenerative diseases with siRNA is particular- ly challenging, however, because siRNAs do not cross the blood-brain barrier and must be directly injected into the brain. Unmodified siRNAs directly injected into the brain are rapidly degraded or cleared through the cere- brospinal fluid. Typical siRNA formulations (e.g., lipid nanoparticles cholesterol bioconjugates) that improve retention and cellular uptake in other organs show pronounced toxicity when administered in the brain. There is a clear, unmet need for methods that improve neuronal delivery of therapeutic siRNAs and minimize off-target effects, toxicity, and immunogenicity. We have synthesized and evaluated an extensive panel of naturally occurring, neuroactive conjugates for brain delivery (including steroids, dopamine reuptake inhibitors, gangliosides, and polyunsaturated fatty acids). We discovered that docosahexaneoic acid (DHA) is amenable to hsiRNA conjugation and exhibits an optimal hydrophobicity profile for brain retention, distribution, and global Htt mRNA silencing. Following a single, intra- cranial injection, DHA-hsiRNA shows diffuse spread and Htt knockdown in the tissues primarily affected by HD. Moreover, there are no indications of neuronal death or immune system stimulation following administration of doses in great excess of what is required for activity. This level of tolerance and diffusion following a single, direct injection has never been documented for a CNS-directed oligonucleotide therapeutic. The mechanism behind this unprecedented safety is unknown. The goal of this proposal is to investigate the pharmacokinetic and pharmacodynamic (PK/PD) properties of DHA-conjugated oligonucleotides in the brain of Huntington's disease mice. This will be accomplished by determining the mechanism of DHA-hsiRNA uptake and metabo- lism and evaluating efficacy and toxicity in Q140 mice, a transgenic strain expressing 140 CAG repeats. The completion of this proposal will offer new insight into the chemical design of siRNAs for delivery to brain. Successful completion will overcome a significant obstacle to the use of siRNA for the treatment of HD and potentially other autosomal-dominant neurological diseases.